In recent years, a method of immobilizing a number of nucleic acid fragment samples on a smooth substrate and thereby determining the sequence information of these fragments in parallel has been developed and being industrialized.
Analysis using, as a nucleic acid sample to be analyzed, whole genome DNA, whole RNA, or cDNA library which is a reverse transcription product of messenger RNA has been conducted popularly. On the other hand, an analysis method with only a specific site of genome DNA or RNA is widening from the standpoint of saving the cost and time for analysis. In addition, superiority of sequence analysis with an accuracy of even one base has been recognized by narrowing a region on a disease-related gene group or genome and instead of it, increasing redundancy of sequence analysis. For example, there is a report on a specific genomic region closely related to diseases such as cancer, Crohn's disease, and schizophrenia. Cancer is a disease triggered by genomic abnormalities. A region to be subjected to sequence analysis differs depending on the tissue where cancer is developed, stage of the pathological condition, polymorphism which individuals essentially have, or the like. There is however a report on structural abnormalities such as deletion, insertion, and abnormality of copy number in a region extending from tensb to tens kb (Non-patent Document 1). In addition, there is a recent report that also in Chohn's disease which is an unexplained nonspecific inflammatory disease causing inflammatory or ulcers in the whole digestive tract region, deletion of abase region about 20k upstream of an immunity-related GTPase family M (immunity-related GTPase family, M) gene is closely related to the onset (Non-patent Document 2) of this disease. There is also a report on a plurality of gene loci related to the onset of schizophrenia (Non-patent Document 3).
Sequence analysis that limits a base region to be analyzed to a specific gene group, a gene group involved in related pathway, a genomic region including a region upstream or downstream of the related gene, “hot spot”, that is, a base region having a high mutation rate per base, or the like is being recognized as markedly useful also from the standpoint of clinical application.
Different from such a sequence analysis that limits a base region to be analyzed, it is, in the conventional method, the common practice to extract, isolate, and elute a desired sequence fragment group by using a microparticle carrier having a probe for capturing a specific sequence fragment or a microarray having the above-mentioned probe immobilized onto a smooth substrate and thereby preparing a nucleic acid sample solution for analysis, and then conduct sequence analysis by using an existing sequencer. This means that a nucleic acid analysis device is manufactured by preparing a specific nucleic acid sample solution, amplifying using PCR, and immobilizing on a measuring substrate to be used for each sequencer and then sequence analysis is conducted using the sequencer. In these conventional methods, a step of extracting and isolating a specific sequence fragment and a series of pretreatment steps of a sample to be analyzed using a sequencer such as a step of introducing an adapter sequence into the end of a sample fragment through ligation and then conducting a PCR amplification reaction in an emulsion (oil droplet) or on a smooth substrate are completely separated from each other so that the conventional methods require a cumbersome wet treatment.